Urinary Nitrate Research Articles

IDF21-0098 Curcumin prevents renal aggravation by ischemia and reperfusion in diabetes mellitus

Background: Diabetes Mellitus (DM) increases the susceptibility to ischemia and reperfusion (I/R). I/R and DM share common physiopathological mechanisms, like inflammation and oxidation. Curcumin, present in Curcuma longa l., is a natural anti-inflammatory and antioxidant, used as a medicine for the treatment of osteoarticular diseases and usually found as a food, in view of its regulation of practical use, it is considered to be used in the prevention of acute chronic kidney disease. Aim: The aim of this study was to evaluate the effect of previous Curcumin treatment on renal function, hemodynamics and oxidative profile of diabetic rats submitted to I/R. Method: Wistar male rats (250–290 g), randomized into four groups: Citrate- received citrate buffer, i.v., on 1st day; DM: received 65 mg/kg streptozotocin (STZ), i.v., diluted in citrate buffer, on 1st day; DM+I/ R: diabetic animals that on 26th day were submitted bilateral renal pedicle clamping for 30 min; DM+I/R+ Curcumin: diabetic animals that received Curcumin 30mg/kg/day, gavage, 17th until the 27th day, and on the 26th day were submitted I/R. The renal function (inulin clearance- InCl, urinary flow- UF, serum creatinine- SCr), renal hemodynamics (renal blood flow -RBF, renal vascular resistance- RVR) and oxidative profile (urinary peroxides- UP, urinary nitrate- UN, thiobarbituric acid reactive substances- TBARS and thiols in renal tissue) were assessed. Results: Curcumin treatment in diabetic animals submitted to I/R insult resulted in increased InCl and decreased SCr, accompanied by increased RBF and decreased RVR with a reduction in UP, UN, TBARS and increase in thiol renal. Discussion: Curcumin prevented renal function decline in diabetic rats submitted to I/R with beneficial repercussions on renal hemodynamics and renal oxidative profile compared with control rats.

The Association Between Environmental Perchlorate, Nitrate, and Thiocyanate Exposure and Oral Pain in NHANES.

AimTo examine the human exposure to perchlorate, nitrate, and thiocyanate, and their associations with oral pain (OP) in the general population from the U.S.MethodsA total of 13,554 participants were enrolled in the National Health and Nutrition Examination Survey. The urinary perchlorate, nitrate, and thiocyanate were measured using ion chromatography coupled with an electrospray tandem mass spectrometry. The multivariable linear and logistic regressions were performed to explore the associations of the urinary perchlorate, nitrate, and thiocyanate, with the prevalence of oral pain. Restricted cubic splines were used to explore the non-linearity.ResultsThere are 3,129 OP cases. There was a higher urinary level of perchlorate, nitrate, and thiocyanate in OP. We found that urinary thiocyanate was positively associated with OP (odds ratio [OR] = 1.06; [1, 1.13]; p = 0.049). Restricted cubic spines revealed that urinary thiocyanate was in a U-shape association with OP.ConclusionsUrinary thiocyanate was in a U-shape association with OP, suggesting that we should keep the exposure of thiocyanate under a reasonable range.

Diet quality and exposure to endocrine-disrupting chemicals among US adults

Human exposure to endocrine-disrupting chemicals (EDCs) may increase risk for chronic disease. Diet is a significant source of EDC exposure, yet healthy diets recommended for chronic disease prevention have not been thoroughly examined for associations with EDC exposure. Using data from the National Health and Nutrition Examination Survey 2013–2016, we examined associations of dietary patterns with exposure to non-persistent EDCs potentially consumed through diet. EDCs were measured in spot urine samples. Diet was assessed using 24-h recalls. Multivariable linear regression was used to examine associations of three healthy diet scores [Healthy Eating Index (HEI), relative Mediterranean Diet (rMED), and Dietary Approaches to Stop Hypertension] and fast-food consumption with EDCs. In fully adjusted models, no diet was associated with exposure to the bisphenols, phthalates, or polycyclic aromatic hydrocarbons examined. A 1-point increase in rMED (of 18 possible points) was associated with 2.7% (95% CI: 1.7%, 3.8%) greater urinary nitrate. A 10-point increase in HEI (of 100 possible points) was associated with 5.3% (95% CI: 2.8%, 7.9%) greater nitrate and 6.8% (95% CI: 4.5%, 9.2%) greater perchlorate. Because perchlorate and nitrate can disrupt thyroid hormone production, we conducted an exploratory analysis to examine whether these chemicals mediate an association between diet and thyroid hormones. A 10-point increase in HEI was associated with 0.6% reduced serum total thyroxine (95% CI: 1.7%, 0.5%) among all adults, with 57.5% of the effect explained by perchlorate. Nitrate mediated an association of rMED with modestly reduced total triiodothyronine among females. Most EDCs examined had no association with the diets evaluated, indicating that recommended healthy diets were not protective against EDC exposures. As observed with two thyroid antagonists, some recommended diets may increase EDC exposures and related adverse health outcomes. Additional work should identify effective food production and processing practices to reduce dietary exposures to potentially harmful EDCs.

Pharmacokinetic Profile of Incremental Oral Doses of Dietary Nitrate in Young and Older Adults: A Crossover Randomized Clinical Trial

BackgroundDietary nitrate consumption can increase concentrations of nitrate and nitrite in blood, saliva, and urine. Whether the change in concentrations is influenced by age is currently unknown. ObjectivesWe aimed to measure changes in nitrate and nitrite concentrations in plasma, urine, and saliva and exhaled NO concentrations after single incremental doses of dietary nitrate in young and older healthy adults. MethodsTwelve young (18–35 y old) and 12 older (60–75 y old) healthy, nonsmoking participants consumed single doses of 100 g, 200 g, 300 g whole beetroot (BR) and 1000 mg potassium nitrate (positive control) ≥7 d apart in a crossover, randomized clinical trial. Plasma nitrate and nitrite concentrations and exhaled NO concentrations were measured over a 5-h period. Salivary nitrate and nitrite concentrations were measured over a 12-h period and urinary nitrate over a 24-h period. Time, intervention, age, and interaction effects were measured with repeated-measures ANOVAs. ResultsDose-dependent increases were seen in plasma, salivary, and urinary nitrate after BR ingestion (all P ≤ 0.002) but there were no differences between age groups at baseline (all P ≥ 0.56) or postintervention (all P ≥ 0.12). Plasma nitrite concentrations were higher in young than older participants at baseline (P = 0.04) and after consumption of 200 g (P = 0.04; +25.7 nmol/L; 95% CI: 0.97, 50.3 nmol/L) and 300 g BR (P = 0.02; +50.3 nmol/L; 95% CI: 8.57, 92.1 nmol/L). Baseline fractional exhaled NO (FeNO) concentrations were higher in the younger group [P = 0.03; +8.60 parts per billion (ppb); 95% CI: 0.80, 16.3 ppb], and rose significantly over the 5-h period, peaking 5 h after KNO3 consumption (39.4 ± 4.5 ppb; P < 0.001); however, changes in FeNO were not influenced by age (P = 0.276). ConclusionsBR is a source of bioavailable dietary nitrate in both young and older adults and can effectively raise nitrite and nitrate concentrations. Lower plasma nitrite and FeNO concentrations were found in older subjects, confirming the impact of ageing on NO bioavailability across different systems.This trial was registered at www.isrctn.com as ISRCTN86706442.

Exposure to perchlorate, nitrate and thiocyanate was associated with the prevalence of cardiovascular diseases

AimsTo determine the association between urinary levels of perchlorate, nitrate and thiocyanate, and the prevalence of cardiovascular diseases (CVD) among general population. MethodsA total of 16, 570 participants were enrolled from the National Health and Nutrition Examination Surveys (NHANES). Urinary levels of perchlorate, nitrate and thiocyanate were measured using ion chromatography coupled with electrospray tandem mass spectrometry. Multivariable linear regressions and logistic regressions were performed to explore the associations of exposure to perchlorate, nitrate and thiocyanate, and the prevalence of total and specific CVD, including chronic heart failure (CHF), coronary heart disease (CHD), angina, heart failure and stroke. Restricted cubic splines were used to explore the nonlinearity. ResultsParticipants with CVD had a lower urinary level of nitrate and thiocyanate (all P < 0.001). A null association between urinary perchlorate and total CVD or specific CVD was observed. Comparing with the lowest quartile, the highest quartile of urinary nitrate was independently associated with a decreased presence of total CVD (odds ratio [OR] 0.66, 95% confidence interval [CI] [0.53, 0.82]), CHF (OR 0.48, 95% CI [0.33, 0.71]), and stroke (OR 0.63, 95%CI [0.45, 0.88]). In addition, per one-fold increasement of urinary nitrate decreased a 0.15-fold prevalence of total CVD, 0.29-fold prevalence of CHF, and 0.16-fold prevalence of stroke. However, for urinary thiocyanate, we found that the 2nd and 3rd quartile were associated with total CVD, the 2nd quartile associated with heart attack, and the 2nd, 3rd and 4th quartile associated with stroke. What’s more, restricted cubic splines confirmed that the relation between urinary nitrate and CVD was linear (P for nonlinearity = 0.242) and the inverse relation between urinary thiocyanate and CVD was nonlinear (P for nonlinearity < 0.001). ConclusionIn the general population, low levels of nitrate were linearly while thiocyanate were nonlinearly associated with an increased presence of cardiovascular diseases.

Effect of curcumin on acute chronic kidney disease due to ischemia-reperfusion syndrome.

To evaluate the effect of curcumin on renal function, hemodynamics, and renal oxidative profile of rats with chronic kidney disease (CKD) subjected to renal ischemia-reperfusion injury (IRI). Wistar rats, 250-300 g, distributed in four groups: Sham (n = 5), CKD simulation; CKD (n = 5), 5/6 renal ablation for CKD induction; CKD + IRI (n = 5), CKD and renal pedicle clamping for 30 minutes; and CKD + IRI+curcumin (n = 5), CKD + IRI, curcumin administration 30 mg/kg/day, orally, for 10 days. Renal function (inulin clearance, urine flow, plasma creatinine), hemodynamics (blood pressure), and oxidative profile (peroxides, TBARS, and urine nitrate, non-protein soluble thiols in renal tissue) were evaluated. The CKD + IRI + curcumin group showed increased inulin clearance and reduced plasma creatinine, decreased RVR and increased RBF, decreased oxidative metabolites in urine and increased thiols in renal tissue when compared with the CKD + IRI group. The treatment with curcumin preserved renal function and hemodynamics of animals with acute CKD, improving oxidative profile, with reduction of oxidants and preservation of antioxidant reserve.

Independent and interactive associations of dietary nitrate and salt intake with blood pressure and cognitive function: a cross-sectional analysis in the InCHIANTI study

Blood pressure (BP) control is a key target for interventions to reduce cognitive decline. This cross-sectional study explored associations between objective (24-hour urine excretion) and subjective (food frequency questionnaire [FFQ]) measures of dietary sodium and nitrate intakes with cognitive function and resting BP in the InCHIANTI cohort. Baseline data from 989 participants aged >50 years were included. In fully adjusted models, participants with concurrent high nitrate and low sodium (Odds Ratio (OR)=0.49, 95%CI 0.32–0.76, p = 0.001) and high nitrate and high sodium (OR = 0.49, 95%CI 0.32–0.77, p = 0.002) 24-hour urinary concentrations had lower odds of high BP than participants with low nitrate and high sodium concentrations. We found no significant associations between sodium and nitrate intakes (24-hour urinary concentrations and FFQ) and poor cognitive performance. Urinary nitrate excretion was associated with lower BP and results appeared to be independent of sodium intake. Further analyses in longitudinal studies are required to substantiate these findings.

The association between environmental exposure to perchlorate, nitrate, and thiocyanate and all-cause and cause-specific mortality.

The aim is to examine the environmental exposure to perchlorate, nitrate, and thiocyanate and their associations with all-cause and cause-specific mortality in a general population. A total of 17,982 participants were enrolled from the National Health and Nutrition Examination Survey and urinary perchlorate, nitrate, and thiocyanate are measured using ion chromatography coupled with electrospray tandem mass spectrometry. Multivariable linear and logistic regression models were performed to explore the associations between urinary perchlorate, nitrate, and thiocyanate with all-cause and cause-specific mortality. Restricted cubic spline models were used to explore the nonlinearity. During a 7.5-year of follow-up, 1730 deaths occurred. Kaplan-Meier analysis suggested that urinary perchlorate, thiocyanate, and nitrate were associated with all-cause mortality (all log-rank P<0.001). After adjusted for traditional risk factors, we found that urinary perchlorate was inversely associated with cancer mortality (hazard ratio [HR] 0.87; 95% confidence interval [0.76, 0.99]; P=0.038) while nitrate was positively related to cancer mortality (HR 1.19, [1.05, 1.34]; P=0.006). Besides, urinary thiocyanate exposure was U-shape associated with the risk of all-cause mortality (P for nonlinearity<0.001). Urinary perchlorate was inversely while nitrate was positively associated with the risk of cancer mortality. The thiocyanate exposure was U-shape associated with the risk of all-cause mortality.

Design and Functionalization of a µPAD for the Enzymatic Determination of Nitrate in Urine.

In this work, the design of a microfluidic paper-based analytical device (μPAD) for the quantification of nitrate in urine samples was described. Nitrate monitoring is highly relevant due to its association to some diseases and health conditions. The nitrate determination was achieved by combining the selectivity of the nitrate reductase enzymatic reaction with the colorimetric detection of nitrite by the well-known Griess reagent. For the optimization of the nitrate determination μPAD, several variables associated with the design and construction of the device were studied. Furthermore, the interference of the urine matrix was evaluated, and stability studies were performed, under different conditions. The developed μPAD enabled us to obtain a limit of detection of 0.04 mM, a limit of quantification of 0.14 mM and a dynamic concentration range of 0.14–1.0 mM. The designed μPAD proved to be stable for 24 h when stored at room temperature in air or vacuum atmosphere, and 60 days when stored in vacuum at −20 °C. The accuracy of the nitrate μPAD measurements was confirmed by analyzing four certified samples (prepared in synthetic urine) and performing recovery studies using urine samples.

Inorganic nitrate attenuates the systemic inflammatory response in typhoid vaccine-induced endothelial dysfunction in healthy volunteers

Abstract Background Inflammatory responses underlie the development of endothelial dysfunction in CVD, however, therapeutics that might target this pathway have not been forthcoming. A key pathogenic mechanism mediating endothelial dysfunction is a reduction in bioavailable (eNOS-derived) nitric oxide (NO). Activation of the non-canonical pathway for in-vivo NO generation might offer an approach to improve NO levels and recover vascular function in pre-clinical models of CVD. Whether this might occur in humans is unknown. Purpose We hypothesize that consumption of inorganic nitrate will lead to increases in bioavailable NO and thus attenuate the inflammatory pathways leading to typhoid vaccine-induced endothelial dysfunction in healthy volunteers. Methods Healthy male volunteers were recruited (n=78) and randomized to receive either beetroot juice containing 8–10mmol nitrate or placebo (nitrate-deplete) juice once daily for 6 days. Participants underwent serial measurements of BP, FMD and GTN-induced brachial artery dilatation, and haematology and biochemistry, before and after typhoid vaccination. Blood, urine and saliva nitrite and nitrate were quantified using ozone chemiluminescence, and leukocyte flow cytometry analysis was conducted. Results 8-hours post-vaccine endothelial function was depressed in placebo-treated volunteers, however this was prevented in nitrate-treated volunteers. This dysfunction was due to impaired endothelial function since responses to GTN were unaffected either by vaccination or dietary intervention (p=0.981). Dietary nitrate resulted in an increase in plasma (p&amp;lt;0.0001), urine (p=0.0006) and saliva (p&amp;lt;0.0001) nitrate, and urine (p=0.0354) and saliva (p&amp;lt;0.0001) nitrite levels. There was a reduction in the proportions of CD14++/CD16+intermediate monocytes in nitrate-treated participants after vaccine (p=0.016, change from baseline between groups). In the nitrate-treated group, less CD14++/CD16+ intermediate monocyte CD62L expression was identified post-vaccine (p=0.0122), compared to placebo, with no difference in soluble plasma CD62L between groups (p=0.875). CD11b median fluorescence intensity was increased in CD3+/CD4+ T-lymphocytes in nitrate-treated volunteers (p=0.0095). Conclusions Dietary nitrate reduced BP, as previously shown, indicating efficacy of the intervention. Importantly, we also now show for the first time that inorganic nitrate suppresses the systemic inflammatory response, specifically by reducing the numbers and activation state of CD14++/CD16+ intermediate monocytes. Furthermore, an increased expression of CD3+/CD4+ T-cell CD11b and preserved FMD in healthy volunteers treated with nitrate, suggests an anti-inflammatory phenotype, induced by the intervention, leading to improved endothelial function. Inorganic dietary nitrate modulates endothelial function through the attenuation of inflammatory responses and may be of potential therapeutic benefit in patients with established CAD. Funding Acknowledgement Type of funding sources: None.

Inorganic nitrate attenuates endothelial dysfunction consequent to systemic inflammation

Abstract Introduction Chronic cardiovascular diseases are characterised by low-grade systemic inflammation and attenuated nitric oxide (NO) bioavailability resulting in endothelial dysfunction. Inorganic nitrate augments NO bioavailability and improves markers of vascular dysfunction in patients with cardiovascular risk factors. However, the exact mechanism of this effect is uncertain. Purpose To determine whether inorganic nitrate supplementation alters systemic inflammation-induced endothelial dysfunction. Methods 62 healthy male volunteers were randomised 1:1 to receive ∼8–10 mmol of dietary inorganic nitrate in beetroot juice or nitrate-free beetroot juice (placebo) once daily for 6 days. Measures of brachial artery flow-mediated dilatation (FMD), brachial blood pressure (BP), pulse wave analysis and carotid-femoral pulse wave velocity (PWV) by Vicorder were taken prior to and at 8 hours after a typhoid vaccine (to induce mild systemic inflammation). Plasma, urine and saliva samples were also collected. Clinicaltrials.gov: NCT02715635. Results Baseline characteristics were similar between the two groups. Inorganic nitrate significantly elevated plasma nitrite (placebo = Δ0.02±0.5 μM, inorganic nitrate = Δ0.63±1.2 μM; p=0.01) and nitrate levels (p&amp;lt;0.0001) compared to placebo. There were significant increases in urine nitrite (p&amp;lt;0.0001) and nitrate (p&amp;lt;0.0001) in addition to salivary nitrite (p&amp;lt;0.0001) and nitrate (p&amp;lt;0.0001) compared to placebo. After 8 hours, typhoid vaccine induced an increase in circulating white cells (placebo = Δ3.34±3.37x109/L, inorganic nitrate = Δ2.9±2.78x109/L; p=0.58) that was similar in in both arms. However, there was a significant reduction in the FMD response in the placebo group at 8-hours post vaccine; an effect that was absent in volunteers treated with inorganic nitrate (placebo = Δ−1.33±1.53%, inorganic nitrate = Δ−0.07±1.84%, p=0.005). Importantly, there were no statistically significant differences in baseline vessel diameter (p=0.78), time to peak diameter in response to flow (p=0.87) and peak shear rate (p=0.57) between the groups. When comparing change from baseline to 8 hours after the vaccine, there were no significant differences in brachial systolic BP (p=0.12), central systolic BP (p=0.12) and PWV (p=0.60) between groups, but a significant reduction in brachial diastolic BP in the inorganic nitrate group (p=0.048). Conclusions Inflammation-induced endothelial dysfunction was prevented in those receiving dietary inorganic nitrate suggesting that elevating circulating nitrite and delivering NO to the blood vessel wall, through dietary approaches may offer potential therapeutic benefit in those cardiovascular diseases which typically exhibit low grade inflammation and deficiencies in bioavailable NO. Funding Acknowledgement Type of funding sources: Public Institution(s). Main funding source(s): British Heart Foundation

Sex-Dependent Effects of Nephron Ift88 Disruption on BP, Renal Function, and Cystogenesis.

Primary cilia regulation of renal function and BP in health and disease is incompletely understood. This study investigated the effect of nephron ciliary loss on renal physiology, BP, and ensuing cystogenesis. Mice underwent doxycycline (DOX)-inducible nephron-specific knockout (KO) of the Ift88 gene at 2 months of age using a Cre-LoxP strategy. BP, kidney function, and renal pathology were studied 2 and 9 months after DOX (Ift88 KO) or vehicle (control). At 2 months post-DOX, male, but not female, Ift88 KO, compared with sex-matched control, mice had reduced BP, enhanced salt-induced natriuresis, increased urinary nitrite and nitrate (NOx) excretion, and increased kidney NOS3 levels, which localized to the outer medulla; the reductions in BP in male mice were prevented by L-NAME. At 9 months post-DOX, male, but not female, Ift88 KO mice had polycystic kidneys, elevated BP, and reduced urinary NOx excretion. No differences were observed in plasma renin concentration, plasma aldosterone, urine vasopressin, or urine PGE2 between Ift88 KO and control mice at 2 or 9 months post-DOX. Nephron cilia disruption in male, but not female, mice (1) reduces BP prior to cyst formation, (2) increases NOx production that may account for the lower BP prior to cyst formation, and (3) induces polycystic kidneys that are associated with hypertension and reduced renal NO production.

Protection of coenzyme Q10 against contrast-induced acute kidney injury in male diabetic rats

BackgroundDiabetes mellitus (DM) is a major risk factor for contrast-induced acute kidney injury (CI-AKI). DM and CI-AKI result in oxidative damage and inflammation that can be reduced when treated with the coenzyme Q-10 (CoQ10). The aim of this study was to investigate the therapeutic potential of CoQ10 in renal function, renal hemodynamics, oxidative profile and renal histology in diabetic rats subjected to CI-AKI.MethodsWistar rats, male, randomized into five groups: citrate: control animals received citrate buffer (streptozotocin vehicle, 0.4 mL); Tween: control animals of CoQ10 treatment received 1% Tween 80 (CoQ10 vehicle, 0.5 mL); DM: animals that received streptozotocin (60 mg/kg); DM + IC: DM animals treated with iodinated contrast (IC, 6 mL/kg); DM + IC + CoQ10: DM animals treated with CoQ10 (10 mg/kg) and that received IC (6 mL/kg). The protocols lasted 4 weeks. An evaluation was made to measure renal function, inulin clearance and serum creatinine, renal hemodynamics by renal blood flow (RBF) and renal vascular resistance (RVR), markers of oxidative stress such as urinary peroxides and nitrate, lipid peroxidation, thiols in renal tissue and renal histological analysis.ResultsDM animals showed reduced renal function, which was followed by an increase inserum creatinine and significant reduction of inulin clearance and RBF. It was noticed an increase in RVR and redox imbalance with higher urinary peroxides and nitrate lipid peroxidation levels with depletion of thiols in renal tissue. IC treatment exacerbated these changes in DM + IC. CoQ10 administration ameliorated renal function, prevented hemodynamic changes and neutralized oxidative damage and progression of the histologic damage in the DM + IC + CoQ10 group.ConclusionThis study demonstrated the renoprotection properties of CoQ10 in an experimental model of risk factor of DM for CI-AKI. CoQ10 presented an antioxidant effect on the CI-AKI in male diabetic rats by improving renal function and renal hemodynamics, preserving morphology and reducing oxidative stress.

Nitrate and nitrite absorption, recycling and retention in tissues of sheep

Dietary nitrate is of increasing interest both for the pharmacological effects of its metabolites as well as its capacity to inhibit methanogenesis in the gut. A sequence of three experiments was conducted to investigate the absorption, metabolism and excretion of nitrate and nitrite through the gastrointestinal tract (GIT) of sheep, and to determine the fate of nitrate and nitrite in body fluids, tissues and faeces after intravenous dosing with 15N-labeled potassium nitrate (K15NO3) and with 15N-labeled sodium nitrite (Na15NO2). In Experiment 1, twelve female Merino sheep were assigned to one of two dietary treatments and adapted to the experimental diet over two weeks. Six sheep were fed a control diet of wheaten chaff mixture (600 g wheaten chaff plus 200 g wheat grain, CON) and six sheep were fed the CON diet with the inclusion of 18 g nitrate/kg DM (Nitrate). After acclimation to the diets, all sheep received a single intravenous dose of K15NO3 and were placed in metabolic cages for daily collection of total faeces and urine over 6 days. Experiment 2 studied movement of an intravenous dose of 15N in body fluids and tissues. Two sheep not adapted to dietary nitrate were dosed intravenously with K15NO3 or Na15NO2 and body fluids and tissue samples were collected 60 min after dosing. Finally, Experiment 3 was conducted to identify and quantify the major sites of nitrate and nitrite transfer within the body, focusing on absorption, partitioning and secretion into the GIT of anaesthetised sheep. A single dose of sodium nitrate (NaNO3) or sodium nitrite (NaNO2) was introduced into the rumen or abomasum or small intestine, and changes in nitrate and nitrite concentrations in other pools, including plasma, urine and saliva, were determined. Results from Experiment 1 showed that urinary recovery of 15N dose in urea after 46 h and total urinary recovery of 15N 141 h after dosing were greater in sheep fed the Nitrate diet relative to CON (P < 0.05). Recoveries of 15N in tissues indicated that nitrate and nitrite principally accumulated in the skin and muscle of sheep (Experiment 2). Finally, Experiment 3 indicated that nitrate and nitrite were rapidly absorbed from the rumen, abomasum and small intestine into the bloodstream. Nitrite was oxidized in plasma and the resultant nitrate was concentrated and recycled via saliva. Appearance of 15N in urinary urea confirmed the passage of plasma nitrate to the digestive tract, via saliva or transruminal flow to be reduced by gut biota to ammonia.

Tadalafil ameliorates bladder overactivity by restoring insulin-activated detrusor relaxation via the bladder mucosal IRS/PI3K/AKT/eNOS pathway in fructose-fed rats

The pathophysiologies of metabolic syndrome (MS) and overactive bladder (OAB) might overlap. Using fructose-fed rats (FFRs) as a rodent model of MS we investigated the effects of tadalafil (a phosphodiesterase type 5 inhibitor) on the dysregulated insulin signalling in the bladder mucosa and bladder overactivity. Micturition behaviour was evaluated. Concentration–response curves on detrusor relaxation to insulin stimulation were examined. Expression and phosphorylation of proteins in the insulin signalling pathway were evaluated by Western blotting. Levels of detrusor cGMP and urinary nitrite and nitrate (NOx) were measured. We observed FFRs exhibited metabolic traits of MS, bladder overactivity, and impaired insulin-activated detrusor relaxation in organ bath study. A high-fructose diet also impeded insulin signalling, reflected by overexpression of IRS1/pIRS1Ser307 and pIRS2Ser731 and downregulation of PI3K/pPI3KTyr508, AKT/pAKTSer473, and eNOS/peNOSSer1177 in the bladder mucosa, alongside decreased urinary NOx and detrusor cGMP levels. Tadalafil treatment restored the reduced level of mucosal peNOS, urinary NOx, and detrusor cGMP, improved the insulin-activated detrusor relaxation, and ameliorated bladder overactivity in FFRs. These results suggest tadalafil may ameliorate MS-associated bladder overactivity by restoring insulin-activated detrusor relaxation via molecular mechanisms that are associated with preservation of IR/IRS/PI3K/AKT/eNOS pathway in the bladder mucosa and cGMP production in the bladder detrusor.

CENTRAL SYSTOLIC BLOOD PRESSURE RELATES INVERSELY TO NITRIC OXIDE SYNTHESIS IN YOUNG BLACK ADULTS: THE AFRICAN-PREDICT STUDY

Objective: Lower nitric oxide bioavailabilty associates with hypertension in diseased and elderly populations. With hypertension known to develop earlier in black populations, we compared both plasma and urinary nitric oxide-related markers and their associations with central systolic blood pressure and arterial stiffness in healthy young black and white adults. Design and method: We included healthy black and white men and women (n = 1110; 20–30 years) and measured central systolic blood pressure and pulse wave velocity, along with both plasma and urinary arginine, homoarginine, asymmetric dimethylarginine, symmetric dimethylarginine, as well as urinary ornithine/citrulline, nitrite and nitrate. Additionally, the urinary nitrate-to-nitrite ratio was calculated. Results: The black men and women had higher central systolic blood pressure and higher plasma arginine and asymmetric dimethylarginine, but lower urinary nitrate and urinary nitrate-to-nitrite ratio (all p&lt; = 0.003) than their white counterparts. In single and forward stepwise multiple regression analyses, we found an inverse association of central systolic blood pressure (adj. R2 = 0.124; β= –0.134; p = 0.006) and plasma homoarginine in black men. Central systolic blood pressure associated inversely with urinary nitrate-to-nitrite ratio in black women only (adj. R2 = 0.171; β= –0.130; p = 0.029). In the white women, central systolic blood pressure associated positively with urinary asymmetric dimethylarginine (adj. R2 = 0.372; β= 0.162; p = 0.015). Pulse wave velocity associated inversely with plasma asymmetric dimethylarginine (adj. R2 = 0.253; β= –0.163; p = 0.024) in the white women only. Conclusions: The lower nitric oxide synthesis and the higher central systolic blood pressure in our black cohort support the notion of a potential increased risk for future large artery stiffness and hypertension development in later life.

Association of exposures to perchlorate, nitrate, and thiocyanate with allergic symptoms: A population-based nationwide cohort study

Allergic diseases have been one of the leading causes of chronic disorders in the United States. Animal studies have suggested that exposures to perchlorate, nitrate, and thiocyanate could induce allergic inflammation. However, the associations have not been examined among general populations. Here, we investigated data of 7030 participants aged ≥6 years from the National Health and Nutritional Examination Survey (NHANES) 2005–2006. Urinary levels of perchlorate, nitrate, and thiocyanate were measured by ion chromatography combined with electrospray tandem mass spectrometry. Information on allergic symptoms (hay fever, allergy, rash, sneeze, wheeze, eczema, and current asthma) was collected by questionnaire. Allergic sensitization was defined by a concentration ≥150 kU/L for total immunoglobulin E (IgE) levels. The associations were estimated using multivariate-adjusted logistic regression models. A positive association was observed for urinary nitrate and eczema (p < 0.001 for the trend). Compared with quartile 1 (lowest quartile), the odds ratios of eczema with 95% confidence intervals [ORs (95% CIs)] from quartiles 2 to 4 were 1.72 (95% CI, 1.41, 2.09), 1.94 (1.53, 2.47) and 2.10 (1.49, 2.97) for urinary nitrate. In addition, urinary thiocyanate was positively related to sneeze (ORQ4 vs. Q1: 1.25, 95% CI: 1.01, 1.55; p = 0.015 for the trend). However, urinary perchlorate was not correlated with any allergic-related outcome. Additionally, the associations were different among subgroups in a four-level polytomous model. Thus, our results suggested that exposures to nitrate and thiocyanate may be associated with allergic symptoms. Further investigations are warranted to concentrate on the practical strategies to monitor exposure levels and the latent mechanisms of the relationship between exposure and allergy.

Effects of processed meat and drinking water nitrate on oral and fecal microbial populations in a controlled feeding study

BackgroundOne mechanism that can explain the link between processed meat consumption and colorectal cancer (CRC) is the production of carcinogenic N-nitroso compounds (NOCs) in the gastrointestinal tract. Oral and gut microbes metabolize ingested proteins (a source of secondary and tertiary amines and amides) and can reduce nitrate to nitrite, generating potentially carcinogenic NOCs. ObjectiveWe evaluated whether nitrate/nitrite in processed meat or water influences the fecal or salivary microbiota. DesignIn this dietary intervention study, 63 volunteers consumed diets high in conventional processed meats for two weeks, switched to diets high in poultry for two weeks, and then consumed phytochemical-enriched conventional processed or low-nitrite processed meat diets for two weeks. During the intervention, they drank water with low nitrate concentrations and consumed a healthy diet with low antioxidants. Then the volunteers drank nitrate-enriched water for 1 week, in combination with one of the four different diets. We measured creatinine-adjusted urinary nitrate levels and characterized the oral and fecal microbiota using 16S rRNA amplicon sequencing. ResultsUsing linear mixed models, we found that, compared to baseline, urinary nitrate levels were reduced during the phytochemical-enriched low-nitrite meat diet (p-value = 0.009) and modestly during the poultry diet (p-value = 0.048). In contrast, urinary nitrate increased after 1-week of drinking nitrate-enriched water (p-value<10−5). Nitrate-enriched water, but not processed meats with or without phytochemicals, altered the saliva microbial population (p-value ≤0.001), and significantly increased abundance of 8 bacterial taxa, especially genus Neisseria and other nitrate-reducing taxa. Meats, phytochemicals and nitrate-enriched water had no significant effects on saliva alpha diversity or any diversity parameter measured for the fecal microbiota. ConclusionThese findings support the hypothesis that drinking high nitrate water increases oral nitrate-reducing bacteria, which likely results in increased NOC. However, meat nitrate/nitrite at the levels tested had no effect on either the gut or oral bacteria. ClinicalTrials.gov IdentifierNCT04138654.

Renal handling of nitrate in women and men with elevated blood pressure.

The inorganic anions nitrate and nitrite are oxidation products of nitric oxide (NO) that have often been used as an index of NO generation. More than just being surrogate markers of NO, nitrate/nitrite can recycle to bioactive NO again. Nitrate is predominantly eliminated via the kidneys; however, there is less knowledge regarding tubular handling. The aim of this study, as part of a large randomized controlled trial, was to explore potential sex differences in renal nitrate handling during low and high dietary nitrate intake. We hypothesized that renal clearance and excretion of nitrate are higher in men compared to women. In prehypertensive and hypertensive individuals (n=231), nitrate and nitrite were measured in plasma and urine at low dietary nitrate intake (baseline) and after 5weeks supplementation with nitrate (300mg potassium nitrate/day) or placebo (300mg potassium chloride/day). Twenty-four hours ambulatory blood pressure recordings and urine collections were conducted. At baseline, plasma nitrate and nitrite, as well as the downstream marker of NO signalling cyclic guanosine monophosphate, were similar in women and men. Approximately 80% of filtered nitrate was spared by the kidneys. Urinary nitrate concentration, amount of nitrate excreted, renal nitrate clearance (Cnitrate ) and fractional excretion of nitrate (FEnitrate ) were lower in women compared to men. No association was observed between plasma nitrate concentrations and glomerular filtration rate (GFR), nor between FEnitrate and GFR in either sex. After 5weeks of nitrate supplementation plasma nitrate and nitrite increased significantly, but blood pressure remained unchanged. FEnitrate increased significantly and the sex difference observed at baseline disappeared. Our findings demonstrate substantial nitrate sparing capacity of the kidneys, which is higher in women compared to men. This suggests higher tubular nitrate reabsorption in women but the underlying mechanism(s) warrants further investigation.

Increase in Bioavailability of Nitric Oxide After Renal Denervation Improves Kidney Function in Sheep With Hypertensive Kidney Disease

[Figure: see text].